False positive reactivity of recombinant, diagnostic, glycoproteins produced in High Five™ insect cells: Effect of glycosylation

Identification, Recombinant Expression, and Characterization of the 100 kDa High Molecular Weight Hymenoptera Venom Allergens Api m 5 and Ves v 3

“…Further support for their relevance in venom allergy stems from the fact that Sf9 insect cells were employed for production of the enzymatically active recombinant proteins. In contrast to High Five cells, Sf9 cells are considered to exhibit significantly reduced a-1,3 core fucosyltransferase activity (39,40), resulting in proteins without immunologically detectable amounts of CCDs (41). Very recently, we were able to demonstrate the validity of these findings in the context of Hymenoptera venom allergy and to develop a strategy for defining the presence or absence of CCD reactivities on insect cell-derived proteins (42).…”

Revealing the anti-HRP epitope in Drosophila and Caenorhabditis

“…In Trichoplusia ni (High Five™) cells, difucosylated glycans are rather common in the overall glycomic profile [48] and are present on various recombinant proteins produced in these cells [49,50]. The presence of core α1-3-fucose in lepidopteran cell lines is not just of significance for what it says about the glycomic potential of insects in general, but is also of interest due to the use of these cells in biotechnology; these aspects are discussed in a couple of recent articles [51,52].…”

Lepidopteran cells, an alternative for the production of recombinant antibodies?

“…While only α1,6 fucose was found in glycoproteins expressed in B. mori and Sf 9 cells, 78,80 α1,3 and α1,6 fucose are found in Masmestra brassicae 78,80 and T. ni (High five TM ) cell lines. 20 The presence of α1,3-linked fucose, a potential allergenic epitope in these cell lines, may constitute a limitation to their use for expressing human glycoproteins. 81 IgG3 or IgM Fc domains can theoretically bind to hexameric C1q molecule, via contact by its CH 2 -CH 3 domain, and activate the classical pathway of complement.…”

“…Fusion proteins such as chimeric-hormone-antibody molecules that achieved in human cells and may even be immunogenic (see below, "Glycosylation in insect cells"). 20 Furthermore, the infectious titers of cell culture supernatant are very low, even at a late time after infection. 19 Therefore Hi-Five cells require infection using a higher titer viral supernatant generated by infecting Sf 9 cells.…”